Thousands of human genes produce multiple distinct mRNA isoforms through alternative cleavage and polyadenylation (APA). APA is often conserved in other mammals. It can dramatically change protein function, e.g., switching between membrane-bound and secreted forms. It can sometimes produce mRNA isoforms that differ in their stability or translation, but we have found that this is uncommon. Instead, we hypothesize that APA is often regulated for the purpose of producing mRNA isoforms that differ in their subcellular localization, such as localization to axons and dendrites of neurons. Our approach to the long-term goal of understanding the regulation and function of APA in mammals is organized around the following specific aims: SA1. Determine the dynamics and function of mRNA 3' ends in neuronal differentiation. We propose to map the dynamics of alternative 3' UTR expression during neuronal differentiation in vitro and to assess the localization properties of alternative 3' UTRs. SA2. Identify factors that control the neuronal program of alternative 3' UTR isoforms. We will integrate data from SA1 and from an ENCODE project related to RNA binding proteins to identify candidate APA regulatory factors, and will then test them by analysis of mRNA isoforms following RNAi, and by use of a metabolic labeling approach to distinguish regulation of cleavage and polyadenylation (CPA) from regulation of mRNA stability. Fundamentally, this proposal seeks to understand the regulation and function of the 3' ends of genes, and to establish their roles in the nervous system, with potential implications for neurological disease and cancer.